IMMUNOLOGICAL REACTIVITY OF ANGIOTENSIN-II RECEPTOR ANTAGONISTS - POSSIBLE IMPLICATIONS FOR RECEPTOR-BINDING SITES

In the present study, we assessed the reactivity with seven anti-angio tensin 11 monoclonal antibodies of three nonpeptide and one peptide co mpounds described as selective antagonists of angiotensin II for AT, ( DuP 753, chloro-5-(hydroxymethyl)-1-[[2'-(1H-tetrazol-5-yl) biphenyl-4 -yl] methyl] imidazole; EXP 3174, 2-n-butyl-4-chloro-5-(carboxylic aci d)-1-[[2'-(1H-tetrazol-5-yl) biphenyl-4-yl] methyl] imidazole) and AT2 receptor sites (CGP42112A, N(alpha)-benzyloxycarbonyl-Arg)Lys-His-Pro -Ile-OH; PD123177, 1-[(4-amino-3-methylphenyl) enyl-acetyl)-4,5,6,7-te trahydro-1H-imidazol[4,5-c] pyridine 6-carboxylic acid), respectively. These studies were undertaken because the reactivity of the monoclona l antibodies with peptide analogs of angiotensin II and the three-dime nsional structure of an angiotensin II-immunoglobulin Fab fragment com plex strongly suggested that the conformations identified by the monoc lonal antibodies were relevant to those involved in receptor binding a s defined by biophysical models supported by structure activity studie s. Surprisingly although three of the compounds were described as comp etitive inhibitors of angiotensin II, binding of the various monoclona l antibodies to either ovalbumin-coupled angiotensin II adsorbed to pl astic wells or I-125-labeled angiotensin II in liquid phase was unaffe cted by any of the nonpeptide antagonists and CGP42112A up to 10(-4) M concentration. The antagonists also failed to bind to rabbit polyclon al anti-angiotensin II antibodies. Direct binding experiments in which solid phase-immobilized angiotensin II and DuP 753 conjugates were in cubated with anti-angiotensin II or anti-DuP 753 monoclonal antibodies , did not show any cross-reactivity. Analysed in the context of most r ecent pharmacological studies of the receptor, these results demonstra ting that the angiotensin II receptor antagonists do not share any epi tope with angiotensin II suggest the presence of two distinct receptor binding sites for the natural ligand angiotensin II and the nonpeptid e antagonists tested. Such an hypothesis is also in agreement with the binding properties of mutated receptors.